Hetero-modification of halloysite nanoclay to immobilize endoinulinase for the preparation of fructooligosaccharides.

Present investigation describes immobilization efficiency of endoinulinase onto hetero-functionalized halloysite nanoclay using 3-aminopropyltriethoxysilane and glutaraldehyde as crosslinkers. Under optimal conditions (APTES 0.75%, sonication time 2.25 h, glutaraldehyde 0.75%, activation-time 65 min, immobilized endoinulinase load 60 IU and coupling-time 1 h), maximum yield in enzyme activity (70.65%) and immobilization (89.61%) was obtained. Developed immobilized biocatalyst shown maximum activity at 65 °C and pH 5.0 with wide range thermal (50-80 °C) and pH (4.0-9.0) stability. Increase in half-life (28.70-fold) of immobilized endoinulinase was observed as compared to free enzyme. An enhanced Km and reduced Vmax of endoinulinase for inulin was recorded after immobilization. Maximum FOSs production 98.42% was obtained, under optimized conditions (inulin 10%; immobilized endoinulinase load 85 IU; hydrolysis-time 10 h and agitation rate 130 rpm) containing kestose (36.26%), nystose (27.02%), fructofuranosylnystose (9.98%) and FOSs DP 5-9 (25.15%). Developed immobilized biocatalyst exhibited a splendid operational stability for 18 batch cycles.

Glutaraldehyde functionalization of halloysite nanoclay enhances immobilization efficacy of endoinulinase for fructooligosaccharides production from inulin.

Current work describes the enhancement of immobilization efficacy of Aspergillus tritici endoinulinase onto halloysite nanoclay using crosslinker glutaraldehyde. Under statistical optimized immobilization conditions, viz. glutaraldehyde 1.50% (v/v), enzyme coupling-time 2.20 h, glutaraldehyde activation-time 1.00 h and endoinulinase load 50 IU, maximum activity yield (65.77%) and immobilization yield (82.45%) was obtained. An enhancement of 1.15- and 1.23-fold in both enzyme activity yield and immobilization yield of endoinulinase was observed, when compared with APTES-functionalized halloysite nanoclay immobilized endoinulinase. Immobilized biocatalyst showed maximum activity at pH 5.0 and temperature 60 °C with broad pH (4.0-8.5) and temperature (50-75 °C) stability. Further, optimal hydrolytic conditions (inulin concentration 8.0%; endoinulinase load 80 IU; agitation 125 rpm and hydrolysis-time 13 h) supported fructooligosaccharides yield (95.44%) in a batch system. HPTLC studies blueprint confirmed 95.44% fructooligosaccharides containing 35.41% kestose, 26.19% nystose and 9.69% fructofuranosylnystose. The developed immobilized biocatalyst shown good stability of 8 cycles for inulin hydrolysis.

Biofuels from inulin-rich feedstocks: A comprehensive review.

Biofuels are considered as a pre-eminent alternate to fossil fuels to meet the demand of future energy supply in a sustainable manner. Conventionally, they are produced from lignocellulosic raw materials. Saccharification of lignocellulosic raw materials for bioethanol production is a cumbersome process as compared to inulin-rich feedstocks. Various inulin-rich feedstocks, viz. jerusalem artichoke, chicory, dahlia, asparagus sp., etc. has also been exploited for the production of biofuels, viz. bioethanol, acetone, butanol, etc. The ubiquitous availability of inulin-rich feedstocks and presence of large amount of inulin makes them a robust substrate for biofuels production. Different strategies, viz. separate hydrolysis and fermentation, simultaneous saccharification and fermentation and consolidated bioprocessing have been explored for the conversion of inulin-rich feedstocks into biofuels. These bioprocess strategies are simple and efficient. The present review elaborates the prospective of inulin-rich feedstocks for biofuels production. Bioprocess strategies exploited for the conversion of inulin-rich feedstocks have also been highlighted.

HPTLC-densitometry quantification of fructooligosaccharides from inulin hydrolysate.

The objective of present research was to develop an easy, precise and accurate HPTLC densitometry method for quantification of fructooligosaccharides (FOSs) from inulin hydrolysate. The chromatographic separation of FOSs was performed on pre-coated silica gel (60, F254) TLC plates using a mobile phase (butanol:ethanol:water, 60:24:16), and densitometry evaluation of FOSs was performed at A500. Both kestose and nystose were successfully resolved with Rf value of 0.43 and 0.34, respectively. The accuracy, reliability and reproducibility of developed method was assessed by percent relative standard deviation of kestose and nystose for instrument precision (1.43% and 1.50%), repeatability (1.48% and 1.56%), intra-day precision (1.60% and 1.63%), inter-day precision (1.62% and 1.66%), limit of detection (4.58 ng/spot and 4.58 ng/spot), limit of quantification (13.87 ng/spot and 13.89 ng/spot) and recovery (98.81% and 98.69%). Moreover, overlapping spectra of test sample with standard confirms the specificity of developed method, which was validated as per ICH guidelines.

Understanding the interactive influence of hydrolytic conditions on biocatalytic production of fructooligosaccharides from inulin.

Statistical optimization of hydrolytic conditions for the production of fructooligosaccharides (FOSs) from pure inulin using Aspergillus tritici endoinulinase was carried out in a batch system. FOSs yield 99.19% was obtained under the optimized hydrolytic conditions i.e. inulin concentration (7.3%), enzyme load (65 IU), hydrolysis time (13 h) and agitation (100 rpm). The closeness of value of co-efficient of determination (R2) to 1, good agreement between model's predicted and experimental values, low percentage error (<5%), high adequate precision (>4%) and F value (11,634.32), and low Lack of fit (0.60) of the designed model authenticates its fitness. High substrate concentration, low enzyme load and short hydrolysis span justifies efficiency of developed process for the preparation of FOSs from inulin using fungal endoinulinase. TLC chromatographic and densitometry studies confirmed the synthesis of short-chain length FOSs. FOSs preparation contained 33.85% GF2 (ketose), 24.50% GF3 (nystose), 7.26% GF4 (fructofuranosylnystose) and 33.58% FOSs of DP5-9.

Purification, thermodynamics and kinetic characterization of fungal endoinulinase for the production of fructooligosaccharides from inulin.

Three-step purification technique (isopropanol precipitation, ion-exchange and size-exclusion chromatography) was used for the purification of an endoinulinase from the culture broth of Aspergillus tritici BGPUP6. The molecular mass of purified endoinulinase was found to be 53.45 kDa and 53.70 kDa by denatured protein gel (SDS-PAGE) and size-exclusion (Sephadex G-100) chromatographic analysis, respectively. Higher Km (0.90 mM), Vmax (19.60 mM/min·mg), Kcat (1.3 × 10-3/min) and Vmax/Km ratio (21.77/min·mg) of purified endoinulinase for inulin than stachyose depicts its higher affinity towards inulin. Purified enzyme was found stable in a pH range 4.0-7.0 with an optimal pH 5.5. The optimal temperature of purified biocatalyst was 55 °C with thermostability in the range of 50-70 °C. D-value and Z-value for endoinulinase at 55 °C was found to be 100.08 h and 11.62 °C, respectively. Thermodynamics inactivation parameters (ΔG, ΔH and ΔS) of endoinulinase shows its wide range thermal stability. Endoinulinase activity was enhanced by CaCl2 and MnSO4, while CuSO4, CoCl2, AgNO3, CdCl2, NiCl2, ZnSO4, BaCl2, HgCl2 and EDTA inhibited the activity of enzyme. Purified endoinulinase was successfully used for the production of fructooligosaccharides from inulin.

Updates on inulinases: Structural aspects and biotechnological applications.

Inulinases are inulin catalyzing enzymes which belongs to glycoside hydrolases (GH) family 32. Bacteria, fungi and yeasts are the potential sources of inulinases. In the present biotechnological era, inulinases are gaining considerable attention, due to their wide range of applications which includes the production of high fructose syrup, fructooligosaccharides and many other important metabolites like bioethanol, organic acids, single cell oil, 2,3-butanediol, single cell proteins, etc. These applications of inulinases have attracted the researchers world-wide to understand the inulin-inulinase interactions for polyfructan hydrolysis. To understand these interactions, the information on structural organization of inulinases is very important which is scarce in literature. The current review highlights the structural and functional properties of inulinases, and difference in their structural organization. The biotechnological potential of inulinases for the production of different bio-products from inulin/inulin-rich raw materials using different bioprocessing strategies has also been elaborated.

Biotechnological applications of inulin-rich feedstocks.

Inulin is a naturally occurring second largest storage polysaccharide with a wide range of applications in pharmaceutical and food industries. It is a robust polysaccharide which consists of a linear chain of ß-2, 1-linked-d-fructofuranose molecules terminated with α-d-glucose moiety at the reducing end. It is present in tubers, bulbs and tuberous roots of more than 36,000 plants belonging to both monocotyledonous and dicotyledonous families. Jerusalem artichoke, chicory, dahlia, asparagus, etc. are important inulin-rich plants. Inulin is a potent substrate and inducer for the production of inulinases. Inulin/inulin-rich feedstocks can be used for the production of fructooligosaccharides and high-fructose syrup. Additionally, inulin-rich feedstocks can also be exploited for the production of other industrially important products like acetone, butanol, bioethanol, single cell proteins, single cell oils, 2, 3-butanediol, sorbitol, mannitol, etc. Current review highlights the biotechnological potential of inulin-rich feedstocks for the production of various industrially important products.

Moral Injury, Spiritual Care and the Role of Chaplains: An Exploratory Scoping Review of Literature and Resources.

This scoping review considered the role of chaplains with regard to 'moral injury'. Moral injury is gaining increasing notoriety. This is due to greater recognition that trauma (in its various forms) can cause much deeper inflictions and afflictions than just physiological or psychological harm, for there may also be wounds affecting the 'soul' that are far more difficult to heal-if at all. As part of a larger research program exploring moral injury, a scoping review of literature and other resources was implemented utilising Arksey and O'Malley's scoping method (Int J Soc Res Methodol 8(1):19-32, 2005) to focus upon moral injury, spirituality (including religion) and chaplaincy. Of the total number of articles and/or resources noting the term 'moral injury' in relation to spiritual/religious issues (n = 482), the results revealed 60 resources that specifically noted moral injury and chaplains (or other similar bestowed title). The majority of these resources were clearly positive about the role (or the potential role) of chaplains with regard to mental health issues and/or moral injury. The World Health Organization International Classification of Diseases: Australian Modification of Health Interventions to the International Statistical Classification of Diseases and related Health problems (10th revision, vol 3-WHO ICD-10-AM, Geneva, 2002), was utilised as a coding framework to classify and identify distinct chaplaincy roles and interventions with regard to assisting people with moral injury. Several recommendations are made concerning moral injury and chaplaincy, most particularly the need for greater research to be conducted.